The Implications of Prostaglandin E2-Wnt Signaling Pathway Interaction in Autism

Background: Exposure to various environmental factors, including infectious and chemical agents, during pregnancy may result in abnormal lipid metabolism and increase the risk of developing Autism Spectrum Disorders (ASDs). Prostaglandin E₂ (PGE₂) is a lipid signaling molecule derived from membrane fatty acids that acts through four EP receptor subtypes, EP1 to EP4. PGE₂ has an important role in brain development and function. Recent studies show an elevated level of prostaglandin metabolites in individuals with autism. Prenatal exposure to a drug misoprostol, a prostaglandin E analogue, during pregnancy has also been linked to ASDs. Our previous studies provided evidence that PGE₂ and misoprostol can elevate calcium amplitude in neuronal growth cones and reduce neurite extensions in a dose-dependent manner. Moreover, literature suggests a cooperative regulation between prostaglandin signaling and early developmental pathways such as Wingless (Wnt).  Overall, the existing literature and our previous data strongly suggest that PGE₂ signaling may play a contributing role in the etiology of ASDs.

Objectives : Our study aimed to determine whether changes in PGE₂ levels affect the function of embryonic neural cell function, to investigate the potential cross-talk between PGE₂ and Wnt signaling pathways, and to elucidate how these molecules may cause abnormal neuronal development.

Methods : Mouse neuroectodermal (NE-4C) stem cells were used as an in vitro experimental model. The study was conducted on undifferentiated NE-4C cells and cells differentiated into neurons and astrocytes. Cells were treated with various concentrations of PGE₂ or misoprostol to determine the effects on gene and protein expression, as well as cell behaviour. Quantitative real-time PCR was used to measure mRNA expression with SYBR Green reagent or Custom TaqMan® Array Plates. Protein analysis was completed using western blot technique. Cell behaviour such as cell movement and growth measurements was recorded using Nikon Eclipse Ti-E microscope with a specialized object tracking module. 

Results : We show that the EP receptor expression changed between undifferentiated and differentiated cells. Relative to undifferentiated NE-4C cells, neurons had a decrease in EP1 and an increase in EP3β and EP3γ expression. Similarly, astrocytes had a decrease in EP1 but an increase in EP3α and EP4 expression. This could indicate that committed and uncommitted cells respond differently to PGE₂. PGE₂ concentration-dependent treatment also regulated the level of EP3α,β,γ and EP4 receptor expression suggesting that exposure to varying doses of PGE₂ could result in differential downstream effects. Interestingly, we also determined that PGE₂ treatment modifies the expression of various Wnt-dependent target genes. Furthermore, PGE₂ appeared to attenuate Wnt-dependent cell behaviour by modifying average velocity, path length, distance, and cell division.

Conclusions : These results indicated that PGE₂ may regulate neuronal stem cell function via activation of EP receptors in a dose-dependent manner and that it may interfere with other key developmental pathways such as the Wnt signaling pathway.  This study expands our knowledge of the important role PGE₂ signaling may have in the developing nervous system and its potential contribution to the pathology of ASDs.